Breast cancers overexpressing HER2/Neu do not respond to anti-hormone therapy and are more aggressive than their non-expressing counterparts. While the use of Herceptin, a humanized monoclonal antibody to HER2/Neu results in significant clinical response, many patients have intrinsic resistance or are expected to develop resistance to this therapy. Understanding the mechanisms by which HER2/Neu induces tumors and regulates their aggressiveness will be critical for developing more effective therapeutics. In this proposal, we describe three aims using cell-based assays and an in vivo transgenic mouse model that are geared towards identifying the regulatory circuitry initiated by HER2/Neu that is ultimately manifest in the formation of aggressive mammary cancer. We will examine three major regulatory components of tumors including the tumor/stroma interface, intracellular signaling targets of HER2/Neu, and a transcriptional nodal point. Regarding the tumor/stroma interface, we found that TGF-(5 signaling is suppressed early during tumorigenic progression due to loss of T-3RI. In contrast, stromally-derived activin appears to activate Smad2 within the tumor periphery, and activin promotes migration of isolated tumor cells. In this proposal, we will determine if activin signaling regulates tumorigenic progression. With regard to intracellular signaling intermediates, we found that rapamycin, an inhibitor of mTOR, causes regression of HER2/Neu tumors without inducing apoptosis or vascular collapse. We will now determine if rapamycin induces excessive autophagy in tumor cells and if tumors rely on a basal level of autophagy for survival. We will also determine if rapamycin regulates metastasis of HER2/Neu tumors and if anti-HER2 antibody therapy potentiates the efficacy of rapamycin. Lastly, we will focus on a transcription regulator, LMO4, which is an intermediate between HER2/Neu and cyclin D1 in breast cancer cells. We will determine if maintenance of LMO4 expression is required for tumor growth as well as identify transcriptional targets of this protein in HER2/Neu-induced tumors. Upon completion of this proposal we will have significantly increased our understanding of the role of activin in tumor-stromal communication, evaluated the potential utility of rapamycin to treat these cancers, and assessed whether LMO4 is an obligate intermediate in tumorigenesis. These studies should reveal additional targets for development of rationally designed therapies for HER2/Neu-induced breast cancer.